Amyloid Formation Of Fish Beta-Parvalbumin Involves Primary Nucleation Triggered By Disulfide-Bridged Protein Dimers

Amyloid formation involves the conversion of soluble protein species to an aggregated state. Amyloid fibrils of beta-parvalbumin, a protein abundant in fish, act as an allergen but also inhibit the in vitro assembly of the Parkinson protein alpha-synuclein. However, the intrinsic aggregation mechanism of beta-parvalbumin has not yet been elucidated. We performed biophysical experiments in combi-nation with mathematical modeling of aggregation kinetics and discovered that the aggregation of beta-parvalbumin is initiated by the formation of dimers stabilized by disulfide bonds and then proceeds via primary nucleation and fibril elongation processes. Dimer formation is accelerated by H2O2 and hindered by reducing agents, resulting in faster and slower aggregation rates, respectively. Purified beta-parvalbumin dimers readily assemble into amyloid fibrils with similar morphology as those formed when starting from monomer solutions. Furthermore, addition of preformed dimers accelerates the aggregation reaction of monomers. Aggregation of purified beta-parvalbumin dimers follows the same kinetic mechanism as that of monomers, implying that the rate-limiting primary nucleus is larger than a dimer and/or involves structural conversion. Our findings demonstrate a folded protein system in which spontaneously formed intermolecular disulfide bonds initiate amyloid fibril formation by recruitment of monomers. This dimer-induced aggregation mechanism may be of relevance for human amyloid diseases in which oxidative stress is often an associated hallmark.